Unknown K. pneumonia and B. cereus

Unknown K. pneumonia and B. cereus

Introduction

To be able to point out the causes of diseases and infections, the world today is very crucial—it where isolation and identification of these causative agents come in. Microbiology is a study of these microorganisms that are responsible for clinical diseases and food poisoning. It is one of the most critical branches of biological sciences that seeks to solve the problems that come as a result of these pathogenic microorganisms. The microorganisms responsible for infections and diseases may be viruses, pathogenic bacteria, pathogenic fungi, or protozoan. Microbiologists need to be able to identify microbes that cause of clinical diseases to give clinical doctors a leeway in the treatment of these particular diseases with a point of reference. Diseases cannot be treated blindly; there is a need for their identification, which will provide the necessary path for a series of drug discoveries. In a clinical laboratory, the two most important approaches to solving medical problems are the isolation and identification of microorganisms. Identification of the microorganisms helps scientists to classify the biodiversity of the microorganism for easy referencing when it comes to mitigation of possible outbreaks of diseases such as polio that have since been completely finished in some developed countries.

Microbiology, as a branch of medicine, has a rich history that has been implicated in infectious diseases. Apart from microorganisms of medical importance, there are those microorganisms of industrial importance, and particularly bacteria and yeast. Microorganisms have been in use in brewing industries, yogurt processing industries, bakeries, among others. It is obvious that not all the bacteria are detrimental; some are referred to as the normal flora. The normal floras are harmless to their hosts, for example, humans and only become harmful when they are compromised. The use of morphological characteristics may not be sufficient enough in the fight against these dreadful microorganisms. The process of isolation and identification of these microorganisms employ the use of molecular approaches and biochemical approaches. Some of the molecular approaches in use may include PCR (Polymerase Chain Reaction) for multiplication of the genes in question, and Gel Electrophoresis for qualitative analysis of the genes in the experiment. The biochemical tests include Coagulase Test, Catalase Test, Nitrate Test, Starch hydrolysis Test, Motility Test, and Gram Staining for microscopy.

In the cultivation of bacteria, different culture media may be used depending on the characteristics of the specific microorganisms to be isolated. The components in the media must include water, energy source, carbon source, and source of nitrogen, mineral salts, and several growth factors. Bacterial cultures are also important when it comes to testing the sensitivity of bacteria to certain antibiotics found in the market today. It is important in keeping track of multidrug resistance posed by some clinically important bacteria such as Staphylococcus aureus. Excessive usage of antibiotics is implicated as a possible cause of bacterial resistance to drugs. The identification and Isolation of such bacteria are key in the quest of combating multidrug resistance by the said bacteria.

In this experimental study, a dichotomous key is employed was used in performing the necessary biochemical tests for the identification of unknown bacteria using the shortest possible route. Using the dichotomous key, it will be possible to identify the two unknown bacteria, #_unknown in the provided suspension of bacterial culture.

Table 1: A number of possible bacteria  in the suspension

Materials and Methods

The experiment in the identification of the two #_unknowns used dwelled on observation of the morphologically and identifiable characteristics, Gram staining techniques, differential and selective culture techniques, and a series of biochemical tests and analysis.

Bacterial culture suspension was provided by the laboratory technician in the lab. Gram staining technique was then performed on the #_Unknown and observable characteristics recorded with reference to the Microbiology Laboratory Manual. The cultivation of the bacteria was done on two plates by inoculating them for the isolation of the colonies and incubated for 24 hours at temperatures of 37°C for one plate and at 30°C for the other plate. It was a means of determining the appropriate temperatures necessary for the growth of the two #_unknowns. The colonies were then obtained after isolating them. One TP (Tryptic soy agar plate) was cultured and incubated for every successive week with for the two organisms. The organism A was cultivated on the right half of the culture plate while on the other hand, the organism B was cultivated on the second half of the culture plate. Another Gram staining procedure was performed on the resultant pure cultures for the observable morphological traits. The microscopy was done using the oil immersion with a bright field at a magnification of 1000x.

The following are plates that were streaked (single) with the bacterial colonies: Tryptic soy agar plate (TP), EMB (Eosin Methylene Blue Plates), and then followed by Mac Conkey Agar plates. Each of the plates in question was made in two halves using a marker. The marker was meant to ensure that the colonies remain in their intended territory on the culture plate. The two halves in each plate were therefore inoculated with microorganisms A and B respectively for the halves. The plates were then incubated at temperatures of 37°C for a period of 48 hours. The growth in the culture plates was halted by the transfer of the pates into a refrigerator until they were ready for use for the rest of the week. The growth patterns for each plate were observed and documented.

The first microorganism to be used in inoculation was B. it was inoculated in Vogues-Proskauer and Methyl red for two days (48 hours) at temperatures of 37°C and then transferred to a test tube. Five drops of methyl red indicator were then added, and the observations recorded. The microorganism, B, was also inoculated into a broth of Mannitol phenol red and incubated at temperatures of 37°C for another period of 48 hours. The procedure was repeated a second time, and the plates transferred into a refrigerator for preservation. The observations were made and documented. A citrate slant test was done on the microorganism B and observations recorded.

For microorganism A, Urease broth was inoculated with the microorganism, and then incubation was done at 37°C for two days (48 hours), and then the broth transferred to a refrigerator for preservation. One Indole broth and one Citrate slant were inoculated with the microorganism A and incubated at temperatures of 37°C for a period of 48 hours. Both of the broth cultures were transferred into a refrigerator for preservation. Catalase test was also performed for the microorganism A as in the laboratory manual. Kovac’s reagent was also used in the experiment. It was added to the broth culture, and results recorded.

Results

Table 2: Results for Microorganism A

Table 3: Results for Microorganism B